5fsh
Crystal structure of Thermus thermophilus Csm6
Structural highlights
FunctionCSM6_THET8 CRISPR (clustered regularly interspaced short palindromic repeat) is an adaptive immune system that provides protection against mobile genetic elements (viruses, transposable elements and conjugative plasmids). CRISPR clusters contain spacers, sequences complementary to antecedent mobile elements, and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA) (Probable). The type III-A Csm effector complex binds crRNA and acts as a crRNA-guided RNase, DNase and cyclic oligoadenylate synthase; binding of target RNA cognate to the crRNA is required for all activities. This protein is not part of the Csm effector complex (Probable). A single-strand-specific endoribonuclease (ssRNase) producing free 5'-OH (PubMed:26763118). Activity is approximately 1000-fold stimulated by cyclic oligoadenylate (cOA); only cyclic tetraadenylate (cA4) stimulates the ssRNase activity while linear oligoadenylates do not activate the RNase (PubMed:28663439). Another study showed stimulation by linear tetraadenylate at very high concentrations, but did not examine stimulation by cA4 (PubMed:28722012).[1] [2] [3] Publication Abstract from PubMedProkaryotic CRISPR-Cas systems provide an RNA-guided mechanism for genome defense against mobile genetic elements such as viruses and plasmids. In type III-A CRISPR-Cas systems, the RNA-guided multisubunit Csm effector complex targets both single-stranded RNAs and double-stranded DNAs. In addition to the Csm complex, efficient anti-plasmid immunity mediated by type III-A systems also requires the CRISPR-associated protein Csm6. Here we report the crystal structure of Csm6 from Thermus thermophilus and show that the protein is a ssRNA-specific endoribonuclease. The structure reveals a dimeric architecture generated by interactions involving the N-terminal CARF and C-terminal HEPN domains. HEPN domain dimerization leads to the formation of a composite ribonuclease active site. Consistently, mutations of invariant active site residues impair catalytic activity in vitro. We further show that the ribonuclease activity of Csm6 is conserved across orthologs, suggesting that it plays an important functional role in CRISPR-Cas systems. The dimer interface of the CARF domains features a conserved electropositive pocket that may function as a ligand-binding site for allosteric control of ribonuclease activity. Altogether, our work suggests that Csm6 proteins provide an auxiliary RNA-targeting interference mechanism in type III-A CRISPR-Cas systems that operates in conjunction with the RNA- and DNA-targeting endonuclease activities of the Csm effector complex. Structural basis for the endoribonuclease activity of the type III-A CRISPR-associated protein Csm6.,Niewoehner O, Jinek M RNA. 2016 Jan 13. PMID:26763118[4] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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